MTT试验检测EV71 2A蛋白对宿主细胞损伤作用

Host cell injuries caused by 2A^(pro) of EV71 detected by MTT assay

目的在Vero细胞、RD细胞和U87细胞3种细胞系中,研究EV71的2A蛋白对宿主细胞增殖能力的影响及对细胞的损伤作用,揭示2A蛋白在EV71的致病机制中所起的作用。方法将野毒株EV71SDLY 107株(强毒株)、EV71SDLY 1株(弱毒株)和嵌合毒株EV71SDLY 107-2A-1感染Vero细胞、RD细胞、U87细胞后,利用MTT试验测定3种细胞在感染病毒后不同时间点的存活情况,绘制细胞生长曲线,比较细胞增殖程度,并利用单因素方差分析法分析3种细胞在感染不同病毒48h后的存活率;结果 EV71SDLY 107株感染Vero细胞和RD细胞后细胞增殖程度和存活率均低于EV71SDLY 1株(t=41.64,P<0.001;t=33.28,P<0.001);EV71SDLY 107-2A-1嵌合株感染Vero细胞和RD细胞后,细胞增殖程度和存活率均高于亲本病毒EV71SDLY 107株,低于EV71SDLY 1株(t=17.97,P<0.001;t=42.09,P<0.01);EV71SDLY 107-2A-1株感染U87细胞后细胞存活率低于EV71SDLY 1株(t=8.85,P<0.001),但与EV71SDLY 107株感染后细胞存活率无统计学差异(t=0.74,P=0.603);不同细胞感染同株病毒48h后,U87细胞存活率均为最低,Vero细胞存活率均为最高。结论 EV71强毒株的2A蛋白对细胞产生的损伤作用大于弱毒株的2A蛋白,提示2A蛋白在EV71对宿主细胞的作用中发挥功能。

We studied host cell proliferations and injuries caused by 2Apro of EV71 in Vero cells,RD cells and U87 cells and to reveal the role of 2Apro in EV71 pathogenic mechanism.Vero cells,RD cells and U87 cells were infected by wild strains EV71 SDLY 107（EV71fatal strain）,EV71 SDLY 1（EV71mild strain）and chimeric strain EV71 SDLY 107-2A-1,the survival of three cell lines at different time post infected by viruses were detected used by MTT assay,the cell growth curves were drew to compare cell proliferation and the survival rates of three cell lines at 48 hours post infected by different viruses were analyzed by ANOVA.Results showed that cell proliferations and survival rates of Vero cells and RD cells infected by EV71 SDLY 107were lower than EV71 SDLY 1（t=41.64,P〈0.001;t=33.28,P〈0.001）.The proliferations and survival rates of Vero cells and RD cells post-infected by EV71 SDLY 107-2A-1were higher than that in EV71 SDLY 107and lower than that in EV71 SDLY 1（t=17.97,P〈0.001;t=42.09,P〈0.001）.The survival rate of U87 cells infected by EV71 SDLY 107-2A-1was lower than that in EV71 SDLY 1（t=8.85,P〈0.001）and no different than that in EV71 SDLY 107（t=0.74,P=0.603）.U87 cells had the lowest survival rate and Vero cells had the highest survival rate at 48 hpost infected by the same virus strain.The 2Apro of virulent strain caused more severe cell injury than 2Apro of attenuated virus,which reveals 2Apro plays a role in the effect of EV71 on host cells.